Cholesterol-oleate-¹⁴C was incorporated into the LDL by the method of Krieger. Using LDL-¹⁴C as a substrate the hydrolysis of cholesterol-oleate was investigated by the rat arterial wall homogenate. Maximum hydrolysis was observed at around pH 4.5 and the hydrolysis was time dependently increased. Cholesterol oleate hydrolysis in the acetylated LDL-¹⁴C was maximum at around pH 4.5. These results suggest that metabolic pathway was almost the same between nature and denatured LDL in the arterial wall. Furtheremore, cholesterol-oleate hydrolysis in glutaraldehyde, MDA treated, platelet aggregation-treated LDLs was investigated. The relationship between lipid acumulation and denatured LDL was discussed.

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Explants of media obtained from the thoracic aorta of young healthy adult male rabbits were cultured in a DME contained 10% calf serum. Then, effects of hyperlipidemic rabbit serum on the cell and organelle membrane phospholipid were determined with the third generation cells. Technique of thin layer chromatography was applied to determine the composition of various phospholipids of the membranes.
Results: By the supplement of hyperlipidemic rabbit serum, smooth muscle cells were proliferated and following changes were obtained; 1) In the membranes of proliferated cell, percentages of free cholesterol decreased and esterified cholesterol increased and 2) On the phospholipids of membranes, percentages of sphingomyelin decreased and phosphatidyl choline increased. These changes of lipid composition of the membranes seemed to depend on the incubation time.
Discussion and conclusion: Above mentioned results seems to indicate that phospholipid composition of proliferative membranes increased their physiological activities since active phospholipids such as phosphatidyl choline increased and inactive phospholipids such as sphingomyelin decreased. And, it is concluded that cell proliferation by the attack of LDL might be a resistance to develop atherosclerosis.

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Hypothesis, firstly proposed by Rokitansky in 1852 and revived by Duguid in 1946, that atherosclerotic plaques result, at least partly, from organization of fibrin thrombi, is fascinating from view point of blood coagulation, although direct evidences have been lacking. Authors have demonstrated that levels of β-thromboglobulin and platelet factor 4 decreased following administration of antiplatelet agents and that concentration of antithrombin III in plasma lowered following decrease in levels of vitamin K dependent procoagulants by administration of warfarin. These results support Astrup's hypothesis that minimal fibrin deposition and its lysis continuously occurrs on vascular intima in normal subjects. Assuming that these hypotheses are true, hypercoagulability of blood in a broad sence may accelerate deposition of fibrin on vascular wall and may lead to progression of atherosclerosis.
Purpose of this study is to elucidate this problem from investigation of blood constituents which affect removal of activated clotting factors which are most important in formation of fibrin. Two major parameters which affect blood viscosity, namely hematocrit values and fibrinogen content, and two most important inhibitors of activated procoagulants, namely antithrombin III and α₂-macroglobulin, were compared to grade of atherosclerosis of coronary and cerebral arteries classified grossly with naked eyes in serial 537 autopsied cases without cancer or DIC.
Following results were obtained: Hematocrit values were significantly higher in patients with marked atherosclerosis of cerebral artery. There were no statistically significant correlations between levels of inhibitors or fibrinogen and the grade of atherosclerosis, although total antithrombin levels, which were generally lower in males because of higher content of α₂-macroglobulin in females, decreased in females with severe atherosclerosis. However, a 54-years old male with congenital dysfibrinogenemia, in whom delayed convertion rate of fibrinogen into fibrin and impaired platelet aggregability induced thrombin and angiographically marked sclerosis of cerebral artery were demonstrated, had repeated attack of cerebral infarction, although risk factor of arteriosclerosis was lacking but hypertension.
From these results, it is concluded that evidences in favor of the thrombogenic theory of atherosclerosis is insufficient and that mild suppression of blood clotting system is not enough to protect one from arteriosclerosis.

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There exist several theories on pathogenesis of arteriosclerosis, in which thrombogenic theory has been one of the main ones.
In thrombogenic theory, platelet would play an important role both in blood stream and in blood vessel walls. It is commonly recognized that platelet itself does not adhere or aggregate in contact with intact vessel walls, e.g. intact endothelial lines. From the standpoint that platelet aggregation is caused either by the endothelial damages or by hyperaggregability due to chemical or hormonal stimulants, we were going to detect wether the platelet aggregation was induced by chemical or hormonal agents, and wether the endothelial cell is exfoliated even from the intact vessel walls.
(Results and Discussion)
1) Arachidomic acid induced platelet aggregation in vitro, which was inhibited by PGE₁ and PGE₂.
2) 1-methionine and 1-homocysteine induced platelet aggregation, which was inhibited by pyridoxal-phosphate.
3) Polysaccharides such as D-glucosamine, D-galactosamine and DEAE-dextran, accelerated the platelet aggregation induced by ADP, noradrenaline and collagen.
4) Polypeptides such as polylysine, protamine and fibrinogen induced platelet aggregation.
5) We could find the relatively large, flat, and enucleated cells (about 25μm×25μm) in the circulating blood. Although the cell itself might be deprived from venous vessel walls, several reports emphasized that this would correspond to the endothelial cells in arterial vessel walls.
From these findings, we would suppose that the endothelial cells are always exfoliated from vessel walls, and that the chemical or hormonal stimulants to platelets may always irrigate platelet function, which would cause platelet aggregation even in normal vessel walls. The homeostasis between vessel walls and platelet sensibility to stimulants might be the most important factor for preventing arteriosclerosis.

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We have analyzed the low density lipoprotein (LDL) receptor activities of fibroblasts from 4 normal subjects, 5 patients with heterozygous familial hypercholesterolemia (FH) and 9 patients with homozygous FH.
In the cells from 4 normal subjects, the receptor binding, internalization and degradation of ¹²⁵I-LDL were 44±3ng/mg protein (mean±S. E. M.), 386±32ng/mg protein and 1335±214ng/mg protein/6hr, respectively. The cells from 5 heterozygotes showed about 46% of the normal activities of receptor. In the cells from 3 homozygotes, the receptor binding, internalization and degradation of ¹²⁵I-LDL were 0.5±0.3ng/mg protein, 11±5ng/mg protein and 5±3ng/mg protein/6hr, respectively. These homozygotes were thought as receptor-negative type. In the cells from other 6 homozygotes, the receptor binding, internalization and degradation of ¹²⁵I-LDL were 5±1ng/mg protein, 33±4ng/mg protein and 66±20ng/mg protein/6hr, respectively. These homozygotes were thought as receptor-defective type. In these 9 homozygotes, three pairs of siblings were included. One pairs of siblings were classified as receptor-negative and two pairs of siblings were classified as receptor-defective. The receptor-negative phenotypes and receptor-defective phenotypes bled true in individual families.
ML-236B, competitive inhibitor of HMG-CoA reductase, completely inhibited the incorporation of [¹⁴C]acetate into digitonin-precipitable sterols in fibroblasts from normal subjects and heterozygous and homozygous patients with FH with the concentration of 0.5μg/ml. However, at 0.05μg/ml of ML-236B sterol synthesis in fibroblasts from homozygotes was not completely suppressed in contrast to normal and heterozygous cells. Moreover, after preincubation with 0.05μg/ml of ML-236B for 24hr in medium containing lipoproteins sterol synthesis in the cells from receptor-negative homozygote showed 75% of the initial activity compared with that of 25% without preincubation. In the cells from a normal subject and a heterozygote, sterol synthesis was inhibited even after preincubation. These results suggest that 1) the inhibitory effect of ML-236B is overcome in homozygote cells by their high intracellular levels of HMG-CoA reductase and 2) that a higher dose of ML-236B may be required to lower serum cholesterol levels in FH homozygotes than in heterozygotes.

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The incidence of ischemic heart desease (IHD) in familial hypercholesterolemia (FH) was studied in 241 heterozygous and 12 homozygous patients.
In a total of 241 heterozygous patients, 118 were males and 123 females. IHD was observed in 46 and 41% of the males and females. The mean age of 24 males and 38 females with coronary insufficiency, and 30 males and 13 females with myocardial infarction was 49±9, 56±12, 49±11 and 65±6 (Mean±S.D.) years, respectively. The mean age of 5 males and 3 females died from myocardial infarction and 4 males and 2 females died from sudden death was 51±16, 63±1, 47±9 and 75±3 years, respectively. These results suggested that IHD was not only more frequent and precorcious in males with heterozygous FH but also more severe than in females.
There were 6 males and 6 females with homozygous FH. Four patients died from sudden death or heart failure. The mean age of death was 28 years (16-42 years). There was no cerebral or peripheral vascular insufficiency.

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The influence of hepatic triglyceride lipase isolated from human post-heparin plasma by the modified method of Ehnholm et al on the platelet aggregation was studied. 0.5ml of the platelet rich plasma (PRP) or the washed platelet rich suspension (WPRP) obtained by the albumin density gradient separation method of Walsh was incubated with 0.5ml of hepatic triglyceride lipase (HTGL) or the buffer as a control for 5 minutes at 37°C, or additionally for 55 minutes at 28°C (in total 60 minutes). The platelet aggregation was induced by collagen, ADP or thrombin after the incubation.
Results were (1) The reduction of the platelet aggregation in the HTGL group was observed in PRP. (2) The greater reduction of the collagen-induced aggregation was seen after the longer incubation for 60 minutes with HTGL. The difference between the HTGL and the control groups was significant. (3) The platelet aggregation in WPRP inclined to be reduced after the incubation with HTGL, but not so remarkable as in PRP. (4) The contamination of antithrombin III in isolated HTGL was found, but did not influence on the platelet aggregation. (5) The reduction of the platelet aggregation by HTGL was similar to that in the post-heparin PRP.
We concluded from these results that HTGL inhibited the platelet aggregation, probably because of the changes in the lipid and prostaglandin metabolism in platelets, on which the changes in lipid metabolism in the plasma reflected partially.

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Serum apolipoprotein A-I (A-I), A-II (A-II), and HDL-cholesterol (HDL-C) levels were determined in patients on chronic hemodialysis. They were all decreased in those patients as compared with normal persons (A-I, 96±19 vs. 113±21 U/100ml, mean±S. D., p<0.05; A-II, 87±20 vs. 108±13U/100ml, p<0.001; HDL-C, 19±9 vs. 59±8mg/100ml, p<0.001). The decrease of HDL-C level was more pronounced than the decreases of apolipoprotein levels. Consequently, the ratio of HDL-C/A-I and HDL-C/A-II decreased significantly (HDL-C/A-I, 0.19±0.06 vs. 0.48±0.06, p<0.001; HDL-C/A-II, 0.22±0.08 vs. 0.53±0.10, p<0.001). No significant changes were observed in the ratio of A-I/A-II. These results suggested qualitative. abnormalities in plasma high-density lipoprotein in patients on chronic hemodialysis.

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Serum HDL₂ and HDL₂-cholesterol levels were measured in order to clarify the metabolism of HDL subfractions. The measurement of HDL₂-and HDL₂-cholesterol was done with high performance liquid chromatography, using the columns for gel permeation chromatography. Enzymatic reaction method was used to measure cholesterol of HDL subfaction.
According to the serum level of HDL-C, 119 persons-21 normal, 67 diabetics, 22 coronary heart disease, 7cerebral thrombosis, and 2 hyperlipidemia-were divided into 11 classes (Fig. 1). Fig. 1 shows that the values of HDL₂-C on the left and HDL₃-C on the right were expressed as slanted bars at each class of HDL-C values, ranged from 25mg/dl or less to 80mg/dl or more. HDL₂-C levels have a tendency to decrease parelleled to HDL-C levels. On the other hand there were no apparent changes in HDL₃-C except under the range of less concentration of 30mg/dl of HDL-C. But in the range of less than 30mg/dl of HDL-C, HDL₃-C also showed its decrease in accordance with the decreasing value of HDL-C. HDL₃-C in the classes ranged 25 to 30mg/dl and less than 25mg/dl of HDL-C revealed significantly lower serum levels than that ranged 30 to 35mg/dl of HDL-C.
The correlations between HDL₂-C levels or HDL₃-C levels and low HDL-C (less than 30mg/dl) levels or high HDL₃-C (more than 70mg/dl) levels were observed (Table 1).
There was significant correlation between low HDL-C and HDL₃-C (r=0.625), and also between high HDL-C and HDL₂-C (r=0.682).
In this study there were none the normal subjects revealed low HDL-C levels.
Considering above results and our former reported observations that HDL-C level depended on HDL₂-C level in the normal subjects, or correlation between HDL₃-C level and HDL-C level in atherosclerotic disease was found, while correlation between HDL₃-C level and HDL-C level was not found in the normal subjects, it should be concluded that HDL₂-C levels are more changeable than HDL₃-C levels in normal subjects, while in atherosclerotic disease HDL₃-C levels are more changeable than HDL₂-C levels.
Therefore, the levels of HDL₃-C may reflect the artheriosclerotic state more than the levels of HDL₂-C.

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Serum total cholesterol (TCH), triglyceride (TG) and HDL-cholesterol (HDL-CH) were measured in 971 healthy Japanese male and LDL-cholesterol (LDL-CH) was calculated by Fried-wald's method. Atherogenic index (AI) was calculated as follows; AI₁=(TCH-HDL-CH)/HDL-CH, AI₂=LDL-CH/HDL-CH. Obesity index was calculated from body height and weight. Drinking, smoking and sporting habits were evaluated individually.
1) The influence of obesity: TG and AI₁ were correlated with obesity index (r=0.376 and 0.210) and HDL-CH was negatively correlated (r=-0.181). There was no good correlation between obesity and TCH, LDL-CH or AI₂.
2) The influence of drinking habit: TG, HDL-CH and AI were higher and LDL-CH was lower in the subjects with heavier drinking habit, especially in the subjects drinking more than 60-80 grams of alcohol several times a week or almost daily. No correlation was found between TCH and drinking habit. In the subjects who quitted alcohol intake, the average level of TG and HDL-CH were close to those of the drinkers, and the level of LDL-CH and AI were close to those of the nondrinkers.
3) The influence of smoking habit: HDL-CH was higher and AI was lower in the non-smokers. No correlation was found between the amount of smoking and HDL-CH or AI. There was no apparent correlation between smoking and TCH, TG or LDL-CH In the subjects who quitted smoking, HDL-CH and AI were close to those of the non-smokers.
4) The influence of sporting habit: TG and AI were lower and HDL-CH was higher in the subjects with sporting habit.
It was concluded that obesity, drinking, smoking and sporting habits might influence the serum level of HDL-CH and atherogenic index by unknown mechanism.

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INTRODUCTION AND METHOD
Clinical significarices of plasma high density lipoprotein (HDL)-cholesterol (Ch) levels have been investigated extensively. However, following questions still remain to be answered:
1) Should all types of hypo-high density lipoproteinemia be treated?
2) Is hypo-high density lipoproteinemia a cuase or result of arteriosclerotic vascular diseases?
3) What kind of subfraction or component is responsible for antiatherogenecity of HDL?
4) Is it possible to prevent or regress atherosclerosis, when hypo-high density lipoproteinemia is recovered for a long time?
We have reported the heterogeneity of hypo-high density lipoproteinemia based on the characteristics of plasma lipoprotein compositions. We proposed to subdivide hypo-high density lipoproteinemia into two groups: Type I and Type II. In hypo-high density lipoproteinemia Type I, HDL-Ch/LDL-Ch ratio is low, and in Type II, it is normal or high. Furthermore, there may be two subtypes in Type I: Type Ia and Ib. In Type Ia, LDL-Ch level is not high, and in Type Ib, its level is high. The purpose of the present study was to investigate further the characteristics of plasma lipoprotein compositions of hypo-high density lipoproteinemia Type II. Our previous study indicated that Type II hypo-high density lipoproteinemia include dyslipoproteinemia observed in thyroid disease, especially hyperthyroidism and liver cirrhosis. Therefore, in the present study, we have investigated the plasma lipoprotein compositions of the patient with hyper- and hypothyroidism and liver cirrhosis. Lipoprotein fractions were isolated by sequential ultracentrifugation as previously reported.
RESULTS
(1) Plasma lipoprotein composition of the patients with hyperthyroidism
Plasma Ch (122±32mg/100ml) and LDL-Ch (68±23mg/100ml) of the patients with hyperthyroidism were significantly lower than plasma Ch (180±36mg/100ml) and LDL-Ch (110±29mg/100ml) in controls. HDL-Ch levels (46±11mg/100ml) was lower than that (54±10mg/100ml) in controls without statistical significance. HDL-Ch/LDL-Ch ratio of the patients was significantly higher than that in controls. Plasma triglyceride (TG) levels were significantly lower than that in controls. TG levels in VLDL, LDL and HDL were as high as those in controls. Plasma and LDL-phospholipid (PL) were significantly lower than those in controls. These alterations in lipoprotein composition were recovered to the values comparable to the values in controls by treatment of hyperthyroidism. Hypo-high density lipoproteinemia (HDL-Ch<40mg/100ml) was observed in 2 of 7 patients and types of hypo-high density lipoproteinemia of two patients were Type II.
(2) Plasma lipoprotein composition of the patients with hypothyroidism
Plasma Ch (264±64mg/100ml) and LDL-Ch (190±54mg/100ml) of the patients were significantly higher than plasma Ch (190±40mg/100ml) and LDL-Ch (119±31mg/100ml) in controls. HDL-Ch (49±15mg/100ml) of the patient was slightly lower than that (52±10mg/100ml) in controls without statistical significance. HDL-Ch/LDL-Ch ratio of the patients was significantly lower than that of controls. LDL-TG, LDL-PL of the patients were increased significantly. HDL-Ch level of one of 4 patients was 38mg/100ml and his type of hypo-high density lipoproteinemia was Type II.
(3) Correlation between plasma levels of T₃, T₄ and TSH and plasma lipoprotein concentrations
Plasma T₃ and T₄ levels showed negative correlations to lipids concentrations in LDL. Plasma TSH concentrations showed positive correlations to plasma, VLDL-and LDL-TG levels.
(4) Plasma lipoprotein composition of the patients with liver cirrhosis
Concentrations of plasma Ch, LDL-Ch, VLDL-TG, plasma PL and LDL-PL in the patients were significa

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Gradient gel electrophoresis (GGE) was used to study molecular properties of HDL subfractions. The principles of this technique is the molecular sieving utilizing a slab of continuous concave gradient (4 to 30%) of polyacrylamide gel. HDL₂ (1.063<d<1.125g/ml) and HDL₃ (1.125<d<1.210g/ml) prepared by sequential ultracentrifugation, were applied to GGE. HDL₃ moved faster than HDL₂ and the ranges of their apparent molecular weight were (1.2-2.9)×10⁵ and (1.9-3.3)×10⁵, respectively. HDL represents a heterogeneous class of plasma lipoproteins not only in terms of flotation properties but also in terms of their metabolic functions. In order to subfractionate HDL in connection with its metabolic functions, we subfractionated HDL₂ by heparin-Sepharose affinity chromatography according to the method of Weisgraber K.H., which could isolate HDL subclasses depending on the presence or absence of Apo E. Fractions of HDL₂ by heparin-Sepharose affinity chromatography were investigated by GGE. The second peak eluted by 0.095M NaCl which was considered as HDL₂ with Apo E moved slower than the first peak eluted by 0.05M NaCl, which was considered as HDL₂ without Apo E, indicating that HDL₂ with Apo E consists of HDL₂ with the larger molecular size. The apparent molecular weight of HDL₂ with Apo E and without Apo E were (1.8-4.6)×10⁵ and (1.0-3.3)×10⁵, respectively. The present study indicated that GGE was simple, rapid and useful technique to evaluate the molecular properties of HDL subclasses.

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It has been known that desquamation of endothelial cell is a factor of atherogenesis. In the present study, we observed desquamated circulating endothelial cells in man by Hladovec's method. Following results were obteined.
(1) The average of number of circulating endothelial cells was 2.79cells/9μl PRP in men, and 2.74cells/9μl PRP in women.
(2) The diameter of these cells is about 20-50μ in average.
(3) The circulating endothelial cells shows as a rule polygonal shape, often with folded or rolled suggesting a marked flatness.
(4) The membrane of cell was stained by Ruthenium Red.
The results imply that endothelial cells exist in blood stream.

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HDL cholesterol is one of the protective markers against atherosclerosis, whereas the cholesterol ratio (plasma total cholesterol minus HDL cholesterol/HDL cholesterol) is known as a risk marker. In this study the effects of genetic and environmental factors were studied on the markers in two groups of twins, young and adult.
In young twins (29pairs, average age=16years), both levels of HDL cholesterol and cholesterol ratio were significantly different between fraternal twins (19pairs) and identical twins (10pairs) (p<0.005). In adult twins (22pairs, average age=33years), there was no difference in the markers between identical twins (16pairs) and fraternal twins (6pairs). With respect to the intrapair difference of HDL cholesterol, adults showed a significantly higher difference than young twins. In identical twins, significant differences of cholesterol ratio and obesity index were observed between young and adult twin groups (p<0.01 and p<0.05, respectively).
These results suggest that levels of HDL cholesterol and cholesterol ratio are under the genetic control during young ages, but that the environmental factors overcome the genetic factors by the adult age. Obesity is influenced predominantly by the environments.

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Serum lipoproteins of diabetic patients who had ever been suffered from cerebrovascular disorders were analyzed by chemical method and preparative ultracentrifugation. The average age of 40 subjects (17 males and 23 females) was 71 years old. Patients of cerebrovascular disorders were consisted of 36 cases of cerebral infarction, 1 case of cerebral hemorrhage and 2 cases of transient ischemic attack.
Clinical parameters were ECG, obesity, diabetic retinopathy and proteinuria. Evaluation of ECG records was done according to Minnesota Code by which ECG abnormalities were classified into 4 groups in terms of ST-T changes, abnormal Q, atrioventricular block, arrhythmia and so on. Degree of obesity was categorized into 3 groups by “obesity index” which was calculated by the formula: [body weight/(body length-100)×0.9]×100. Grades of diabetic retinopathy were divided into 3 groups according to Scott's criteria.
On the other hand, laboratory data were assembled about HDL-cholesterol, lecithin-cholesterol acyltransferase (LCAT) and malondialdehyde (MDA). Subfractions of serum lipoprotein were separated into VLDL, LDL, HDL₂ and HDL₃ by preparative ultracentrifugation.
Concentration of apolipoprotein AT (Apo-AI) in HDL₂ and HDL₃ was determined by radial immunodiffusion, and amount of protein moiety in HDL₂ and HDL₃ was measured by the method of Lowry et al.
Interrelationship between above-mentioned clinical parameters and lipoproteins was investigated. Correlations between fractions of lipoproteins were estimated.
As to results, no significant correlations were found between MDA and any clinical parameter or any lipoprotein fraction.
Percentage of HDL₃-trigriceride was high both in the group of advanced stages of diabetic retinopathy and in the group of proteinuria. Therefore, it is presumed that HDL₃-triglyceride may be concerned with progression of diabetic microangiopathies.
The most remarkable result was the interrelationship between abnormalities of both ECG and lipid metabolism: HDL₂-cholesterol, HDL₂-phospholipid and HDL₂-Apo-AI were significantly lower in the ECG group of which abnormalities was most marked than in the other groups of lesser abnormalities. This suggests that reduction of HDL₂ subfraction increases the incidence of ischemic heart disease.
So-called atherogenic index which is the ratio of [total cholesterol minus HDL-cholesterol] to HDL-cholesterol showed a strong inverse correlation with cholesterol and phospholipid of HDL₂ subfraction. This data supports the proposal that the lipoprotein most related to atherosclerosis is HDL₂ subfraction and increase of HDL₂-phospholipid as well as HDL₂-cholesterol plays an important role in prevention against atherosclerosis and ischemic heart disease.

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The following experiments were performed in 37 cases of ischemic heart disease (IHD) and 11 cases of healthy persons (HP) to investigate the pathogenesis of IHD. Each serum lipoprotein such as VLDL, LDL, HDL₂₊₃ and VHDL which were separated by ultracentrifugation, was dialysed into running tap water by using the Kanazawa's method and was fractionated into Ultrawatersoluble lipoprotein (UWS-LP) and non-water-soluble lipoprotein (non-WS-LP). In IHD and HP the concentrations of Cholesterol (CH), triglycerides (TG), phospholipid (PL) and binding hexose (BH) contained in those lipoproteins were compared.
1) Serum total lipids didn't show significant differences between IHD and HP, but there were clear differences in UWS-LP and non-WS-LP.
2) By the same comparison, marked differences were found between IHD and HP in lipids concentration of UWS-LP and non-WS-LP from each lipoprotein fractionated by ultracentrifugation. In each fraction of VLDL-CH, LDL-CH and HDL₂₊₃-CH, they were remarkable.
3) Both groups showed significant difference from each other in VHDL-BH.
4) A study from both the viewpoint of lipids and binding hexose is neccessary to investigate the pathogenesis of ischemic heart disease. And also it is useful to study precisely from the aspect of ultrawatersoluble lipoprotein.

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Lipoprotein lipase (LPL) was purified from bovine milk using heparin-Sepharose 4B column Chromatography. About 60% of LPL activity was decreased for 4hr. at 30°C but the addition of Dentran sulfate (DS) did not decrease the activity.
DS increased the hydrolysis of VLDL-TG by LPL. From the results using BioGel A-5m, DS bound to the VLDL-LPL complex and it was suggested that VLDL-TG hydrolysis was enhanced by the formation of VLDL-LPL-DS complex.

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The efficacy of AZ 1355 (300mg/kg, po) was compared with clofibrate (CPIB) using Triton hyperlipidemic mice. AZ 1355 reduced serum cholesterol and the magnitude of reduction was similar to that of CPIB.
AZ 1355 (50, 100mg/kg) and CPIB (100mg/kg) were given orally for 4 weeks to rats fed on either CE-2 or a lipid-rich diet. When the rats were fed on CE-2, CPIB reduced the serum cholesterol and triglyceride but AZ 1355 did not. But with the lipid-rich diet AZ 1355 was more effective than CPIB.
Both agents (100mg/kg) were given orally for 4 weeks to golden hamsters fed on either CE-2 or a 10% butter diet. With both diets AZ 1355 reduced the serum triglyceride more than CPIB did. The dose-response relationship of AZ 1355 was examined using hamsters. The serum triglyceride was reduced by doses above 25mg/kg. In addition, the highest dose (100mg/kg) elevated HDL-cholesterol.
In the rabbits fed on 1% cholesterol diet for 12 weeks, AZ 1355 at a dose of 80mg/kg inhibited an elevation of the serum cholesterol.
The effects of AZ 1355 and its major metabolite, AZ 1355F (having hypolipidemic activity weaker than that of AZ 1355) on platelet aggregation in vitro, arachidonic acid (AA) shock in rats and the formation of Prostaglandin I₂ (PGI₂) and thromboxane A₂ (TXA₂) were studied. PGI₂ and TXA₂ were determined as 6-ketoPGF_1α and TXB₂, respectively.
AZ 1355 above 15μM partially prevented the aggregation induced by collagen or AA and completely inhibited the aggregation above 250μM. AZ 1355 weakly inhibited the aggregation.
AZ 1355 (po) and AZ 1355F (po, iv) dosedependently protected the rats from stroke death induced by AA. But the protective activity of both agents was weaker than that of aspirin.
AA is converted to TXB₂ as the major product and 6-ketoPGF_1α as the minor product, when incubated a mixture of rat platelets and swine aortic microsomes. AZ 1355 inhibited the TXB₂ synthesis at concentration higher 13μM, but 6-ketoPGF_1α formation was unaltered. AZ 1355F also inhibited TXB₂ synthesis, but accelerated 6-ketoPGF_1α production. AZ 1355 and AZ 1355F exerted activities similar to aspirin and imidazole, respectively. The properties of AZ 1355F concerning TXB₂ and 6-ketoPGF₁ are favorable as an anti-atherosclerotic agent.

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Authors have reported on biochemical actions of active principles, esp. saponins, in panax ginseng. The present study was undertaken to investigate the effects of ginseng principles on lipid metabolism in the rats, as well as clinical effects of red ginseng powder on hyperlipidemic patients. Male rats of Sprague-Dawley strain were used. Plasma cholesterol and triglyceride levels of rats fed a 1% cholesterol-0.5% cholic acid diet for 7 days were both high, but the intramuscular injections of either of ginseng saponins (fraction 4), saikoseponins or glycyrrhizin reduced plasma lipid levels. (Fig. 1)
Radioactivity of cholesterol remained 48 hours after interaperitoneal injection of 4-(¹⁴C) cholesterol. The plasma of rats administered with ginseng saponins was significantly lower than controls. (Table 2)
Fecal excretion of radioactive bile acids and sterols after intraperitoneal injection of 4-(¹⁴C) cholesterol was accelerated by ginseng saponins. (Table 3). These results may suggest that ginseng principles accelerated cholesterol turnover and reduced plasma cholesterol level.
To check the net effect of ginseng, effects of oral administration of red ginseng powder for 90 days on plasma lipids, lipid peroxide, platelet adhesiveness and atherosclerosis were investigated using rats fed a high cholesterol-diet. (Table 4) Red ginseng powder used was manufactured at Office of Monopoly, Republic of Korea. Total cholesterol, triglyceride, NEFA and total cholesterol-HDL-cholesterol/HDL-cholesterol, i.e., so called atherogenic index, were much decreased, while HDL-cholesterol and phospholipid were significantly increased. Lipid peroxide remained unchanged. Platelet adhesiveness was decreased. (Table 5)
Fatty liver induced by high cholesterol-diet feeding was improved by simultaneous oral administration of red ginseng powder. Hepatic total cholesterol and triglyceride were decreased, while phospholipid were increased. (Table 6) Macroscopically color of the liver was quite different between the cholesterol diet-fed and the cholesterol plus ginseng diet-fed, that is, yellow in the former and red in the latter. Histological examinations revealed that fatty infiltration was improved by ginseng administration.
Five normal volunteers and 6 hyperlipidemic patients received 4.5g/day of red ginseng powder manufactured by office of Monopoly, Republic of Korea for a week (Table 7). Serum HDL-cholesterol was elevated and triglyceride was declined, while serum cholesterol and lipoproxide showed no significant change. Over fifty patients of hyperlipidemia are receiving ginseng powder. The data will be published.

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It has been suggested that a balance between prostacyclin and thromboxane A₂ may play an important role in the homeostasis of the circulation and in the genesis of atherosclerosis.
Prostacyclin inhibits platelet aggregation and causes vasodilatation and prostacyclin synthetase is inhibited by lipoperoxides.
The experimental models of atherosclerosis in rabbits were produced by atherosclerotic diets. In these models the aortic prostacyclin generating activities and the plasma lipoperoxide levels were investigated and the effects of niceritrol, an antilipidemic agent, on them were also studied.
Rabbits were killed under ether anesthesia, descending thoracic aortas were rapidly removed and washed in cold saline. The aortas were cut into thin rings and incubated in 1ml of krebs' solution for 10min at 22°C. Prostacyclin released from the aortic rings were estimated in comparison of its antiaggregatory activity on human platelets with that produced by the known amounts of the sythetic prostacyclin.
Following results were obtained.
1) Decreased prostacyclin generating activities were found in the aortic tissues of the atherosclerotic rabbits fed for two months with 0.5% cholesterol containing diet and the changes were reversed by the additional supplement of 0.5% niceritrol in it. The same kind of decreases were seen in the rabbits within 2-4 weeks of 1% cholesterol and 3% olive oil feeding.
2) The elevated plasma lipoperoxide levels were observed in the 0.5% cholesterol fed rabbits and these elevations were restrained by the addition of niceritrol.
3) A good inversed correlationship was shown between aortic prostacyclin generating activities and plasma lipoperoxide levels.

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The relationship between arterial calcification and serum estradiol levels was studied in 77 postmenopausal female and 59 male subjects clinically and then estradiol binding study was performed by using cultured rat aortic smooth muscle cells.
In both sexes, subjects with iliac artery calcification had rather lower serum estradiol levels (8.4±1.4pg/ml in the females, and 19.2±2.5pg/ml in the males) than controls (16.1±1.6pg/ml in the females, and 29.7±2.4pg/ml in the males). The bone mineral content of females with iliac artery calcification (0.44±0.02g/cm²) was lower than controls (0.52±0.01g/cm²); a positive correlation between serum estradiol levels and bone mineral content was found in the females. However, bone mineral content did not significantly differ between males with and without arterial calcification (0.67±0.03g/cm² in the former, and 0.65±0.02g/cm² in the latter). These results indicate that arterial calcification and increased bone resorption are both individual results of estrogen deficiency. So, nextly, to determine whether aortic smooth muscle cells possess estradiol receptors or not, an estradiol binding study was performed in cultured rat aortic smooth muscle cells by using an intact-cell assay. Specific binding for estradiol is shown by the cells, and the binding reaches a plateau after 15min of incubation at 37°C. The binding curve shows a saturable binding process. Scatchard analysis reveals a single population for estradiol binding with a dissociation constant of 5.0×10^-8M and specific binding sites of 64 fmole per 10⁶ cells (38, 500 sites per cell). Molecular specificity shows highly specific binding with estradiol. Among various steroids tested, only testosterone at 10^-5M competes slightly (9.3%) with the estradiol-specific binding. These data indicate the existence of specific, high-affinity and limited-capacity binding sites for estradiol in rat aortic smooth muscle cells. This accords with the concept that estrogen responsiveness in aortic smooth muscle cells is mediated through the estrogen-receptor binding in the cells.

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Serum lipids, platelet lipids, volume, content of ATP and ADP and platelet aggregation were examined in diabetic patients with hyperlipidemia before and after one month treatment with nicomol.
Nicomol decreased very low density lipoprotein and triglyceride from 304mg/dl to 202mg/dl and 282mg/dl to 193mg/dl, respectively. No changes were observed in platelet volume, content levels of ADP and ATP and aggregation induced by ADP and epinephrine after treatment with nicomol, whereas a small amount of cholesterolester, triglyceride, free cholesterol and phosphatidylethanolamine estimated per 10⁸ platelets were decreased. Enhanced platelet aggregation induced by arachidonic acid was observed in the patients whose very low density lipoprotein reduced remarkably.

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Length-tension relationships were obtained in aortic strips from six male atherosclerotic rabbits and compared with those from six age and sex-matched normal rabbits. Diffuse atherosclerotic lesions were produced by selective removal of the vascular endothelium with an intraarterial balloon catheter followed by cholesterol-rich diet for two months.
Serum cholesterol levels of atherosclerotic and normal rabbits were 1421.4±242.5mg/dL, 19.30± 0.60mg/dL respectively. Stretch-passive tension curve and stretch active tension curve developed by KCl 60mM of atherosclerotic specimens significantly shifted to the left. Altered length-tension relationship indicates decrease of elasticity in atherosclerotic aortic strips and are thought to be associated with enhanced production of collagen involved in the atherosclerotic process.

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